Biological RNAs undergo extensive post-transcriptional modifications. These modifications are important in promoting the accuracy and efficiency of gene expression in biological systems. To decipher the functions of RNA modifications, robust analytical tools and methods that enable scientists to monitor, manipulate, and transcriptome-wide mapping RNA modifications are highly needed. In this multi-phase SBIR program, Guild Biosciences Inc. (GBS) proposes to address this issue by: 1) developing a library of affinity antibodies and peptide sequences specific to modified nucleosides by using phase-antibody display libraries and peptide phase-display library, 2) developing a novel immunoassay system- open immunocomplex assay with the developed antibodies and peptides for rapid and quantitative detection of modified nucleotides in RNA samples, and 3) incorporating the immunoprecipitation capability of the developed affinity reagents with next-generation sequencing platforms to develop method for global mapping of RNA modifications with high accuracy. Successful completion of the proposed research will add much needed analytical tools and methods in terms of affinity reagents, immunoassay systems and deep sequencing method, which will significantly expand the tool box for the epigenetic research. Several commercial viable products, such as affinity reagents (antibodies, peptides) recognizes the chemical structures of modified RNA nucleosides, bioassay kits for detection/quantitation of RNA modifications, and sequencing preparation kit for deep RNA sequencing will be generated as the ultimate outcome of this proposed research.